PROJECT SUMMARY/ABSTRACT - Project 2 Although controversial, epidemiologic studies have indicated that conception through assisted reproductive technologies (ART), such as in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI), presents a risk for autism spectrum disorders (ASD) and other neurological defects. ART has also been implicated in mediating long-term epigenetic dysregulation, thereby contributing to developmental diseases in humans. To date, genetic variance accounts for fewer than 10% of examined ASD cases, reinforcing the notion that environmental and epigenetic factors underlie ASD incidence. Indeed, culture conditions used in current ART procedures appear to modify epigenetic patterns of normal embryonic development. Additionally, we have identified a potential candidate gene, Shank3, whose genetic and/or epigenetic disruption associates with ASD-like phenotypes in mice. However, the extent to which epigenetic dysregulation directly underlies the ART-associated increased risk for ASD remains undetermined. This is in part due to the lack of a gene-by- environment model relevant to ASD or other neurological disorders. To address this, massively parallel high- throughput sequencing technologies will be employed to examine and characterize the DNA methylomes and gene transcriptomes of brain from IVF-, ICSI-, and naturally-conceived mice (Aim 1). Additionally, comprehensive behavioral assessments will be performed on all mice to link observed epigenetic alterations with social and/or cognitive impairments that characterize ASD (Aim 2). By integrating the genome-wide datasets with behavioral information, candidate ART-associated biomarker genes that are epigenetically dysregulated, including Shank3, will be identified and further investigated during the course of embryonic and brain development (Aim 3). Collectively, addressing these proposed aims will yield a detailed, temporally- defined gene network that underscores the relationship between the pre-implantation environment and the epigenetic mechanisms that play critical roles in neurodevelopment, and help establish a gene-by-environment model of ASD for future investigation to better understand its etiology. Importantly, data from this study will lay the foundation for objectively and rationally altering current ART methodologies to reduce the penetrance of ART-associated ASD and related neurological defects in humans. This work is therefore consistent with the mission of NIH/NIGMS in that it will add to the fundamental knowledge about how the environment of early development may alter normal epigenetic processes that adversely impact long-term health and establishes a novel model that can be applied to inform clinical practices.